In a splitroot system, the influence of mycorrhization of tomato plants with the vesicular-arbuscular mycorrhizal fungus Glomus intraradices on physiology and shikimate pathway transcription was investigated to distinguish between local effects in the mycorrhizal roots and systemic effects in the shoot and in the non-mycorrhizal part of a half-mycorrhizal root. Mycorrhization caused a growth depression and reduced concentrations of elemental carbon and carbohydrates in mycorrhizal and half mycorrhizal roots compared to controls. The two parts of the half mycorrhizal root showed the same low carbon concentration, indicating a systemic effect on carbon availability in the root and the great sink strength of the fungus. Despite, in a developed symbiosis the elevated nitrogen concentration in shoots and roots of mycorrhizal plants, with higher concentrations in the mycorrhizal part of the half mycorrhizal roots, indicated a better supply of mycorrhizal roots and shoots with nutrients, on the cost of nitrogen supply of the non-mycorrhizal part of the root. Although increased nitrogen levels could lead to increased amino acid synthesis, the biosynthesis pathway for the three aromatic amino acids, the shikimate pathway, was not regulated in this later stage of the symbiosis. However, elevated shikimate pathway transcripts in mycorrhizal roots in the early stage of the symbiosis were demonstrated for the first time. This indicates an involvement of the shikimate pathway in early defence responses against the fungus and an influence of changes in carbon status and sugar metabolism on the pathway. A more detailed look to the entry enzyme of the shikimate pathway in plants revealed that one of its two isoforms (DAHPS2) was upregulated by mycorrhization. This one was also induced by short-term ozone exposure, whereas the other was unaffected under the investigated conditions. Furthermore, an influence of mycorrhization on the shoot reaction to ozone was found. Dependent on the mycorrhization rate, an additional treatment with ozone caused additive DAHPS induction of the second isoform in shoots. VOC emissions and glutathione concentrations were only elevated in shoots of non-mycorrhizal plants after ozone exposure, indicating changes in root-shoot interactions involving signalling cascades. Neither early jasmonic acid or hexenal induction nor later methyl-salicylate emissions seem to be relevant in the regulation of DAHPS in response to ozone. Moreover, ozone alone did not only induce the shikimate pathway in shoots, but there was also an isoform specific induction of DAHPS transcripts in roots after ozone treatment, what would require a fast transduction of a shoot signal to the roots. Whether the signalling from shoot to root after ozone exposure is mediated by the same compounds as the root to shoot signalling in the mycorrhizal symbiosis still remains unclear. Furthermore, the different affected pathways and substances may be influenced by different signalling cascades, reflecting the various re-programming in plant metabolism during interactions with belowground symbionts and aboveground environmental parameters. 1 Zusammenfassung ___________________________________________________________________________ Zusammenfassung

Dynamics of leaf and root growth is dependent on endogenous control and environmental impact. Both factors influence carbon partitioning, because a coordinated carbon flux is necessary for maintenance of growth. The resulting carbon allocations are controlled by both sink demand and source control of photosynthate production (Andersen, 2003). Partitioning between the competing sinks is determined by the relative sink strength, which is influenced by abiotic and biotic factors (Biemelt & Sonnewald, 2006). Thus, coordination between root and shoot is necessary to control the carbon partitioning and nutrient acquisition. CO 2assimilation is dependent on the nitrogen supply of the shoot (Khamis et al., 1990) and the nitrate uptake is dependent on a continuous flow of carbohydrates to the root (Rufty et al., 1981). Assimilation of nitrogen and sulphur includes reactions that are among the most energy-requiring reactions in living organisms and thus are strongly regulated at several levels (Taiz & Zeiger, 2002). Source-sink interactions are not only important for normal growth and development, but may also play a role in plant-microbe interactions (reviewed in Biemelt & Sonnewald, 2006). Not only plant pathogens such as bacteria, fungi or viruses, but also mycorrhizal fungi evolved strategies to change plant metabolism to their own benefit. The mycorrhiza symbiosis provides a great carbon sink in roots and therefore has impact on shoot carbon metabolism and the balance between carbon flow into primary and secondary metabolism. Ozone is another factor varying C allocation. Current levels of ozone are capable of altering the timing and quantity of carbon flux to soils (Andersen, 2003), affecting interactions with the rhizosphere and thus root associated microorganisms and symbionts. Short ozone pulses could result in a short-term export stop of carbohydrates and induce accumulation of N-rich secondary compounds. The shikimate pathway is a pathway which could ensure the described coordination between carbon and nitrogen metabolism, which allows the plant to react to varying environmental conditions as it leads to the synthesis of nitrogen rich compounds as well as phenylpropanoids and other carbon-rich secondary metabolites, which lack nitrogen (Coruzzi & Bush, 2001; Walch-Liu et al., 2005).

1.1 The mycorrhiza provides a strong C-sink in roots Mycorrhiza is a very old symbiosis between soil-borne fungi and the roots of higher plants. Plants benefit from the symbiosis by a better supply with nutrients like phosphate and nitrogen, while the fungus is supplied with carbon by the plant. This symbiosis has a strong influence on the plant metabolism and its carbon-nutrient-balance. The first bryophyte-like land plants in the early Devonian (400 million years ago) had already endophytic 3Introduction ___________________________________________________________________________ associations resembling vesicular-arbuscular mycorrhiza (VAM). It is suggested that these mycorrhizal fungi assisted in their colonisation of land (reviewed by Brundrett, 2002; Harrison, 2005). These associations occur in terrestrial ecosystems throughout the world and have a global impact on plant phosphorus nutrition. Trappe (1994) defined mycorrhizas as “dual organs of absorption formed, when symbiotic fungi inhabit healthy absorbing organs (roots, rhizomes or thalli) of most terrestrial plants and many aquatics and epiphytes”. He also suggested that mutualistic functioning of these associations should be a defining criterion of the term mycorrhiza, which was first mentioned to the peculiar association between tree root and ectomycorrhizal fungi (Frank, 1885). The VAM fungi are obligate biotrophs and depend on the plant for supply of carbon. Until now VAM have been found in a wide range of habitats (Strack et al., 2003), mainly in the roots of angiosperms, gymnosperms and pteridophytes but also in some mosses and lycopods (Smith & Read, 1997).

On the basis of morphological criteria, how the fungal mycelium relates to root structures, there are two major mycorrhizal groups: the endomycorrhizas and the ectomycorrhizas. The endomycorrhizas are again subdivided into three groups: the ericoid, the orchidaceous and the VAM. The VAM fungi belong to 6 genera and were first described by Nägeli (1842). Their taxonomy is based mainly on morphological characteristics of the spores. Glomus is thought to be the most abundant genus among soil fungi (Marschner, 1995) and was first described by Tulasne & Tulasne (1844). VAM are characterised by formation of branched haustorial structures (arbuscules) within the cortex cells of the plant and by a mycelium which extends into the surrounding soil. In crop plants, arbuscules, the major site of nutrient exchange between fungus and host plant, are short-lived structures in the root cortex, which senesce 3-4 days after production (Bonfate-Fasolo, 1986). This means that new arbuscules are formed throughout the whole symbiosis. In addition, many but not all VAM form lipid-rich storage organs (vesicles) within the plant roots.

The first published experimental mycorrhization of tomato was possibly done by Mosse (1956) with Endogone in open pot experiments. This species was re-named Glomus mosseae later. Another species often used in experiments on VAM is Glomus intraradices, first isolated in Florida and described by Schenck & Smith (1982). It is well known from many experiments with different fungi and plants that a given VAM fungus may have totally different effects depending on the affected plant species (van der Heijden et al., 1998). In general, the plant growth response to VAM colonisation depends on the balance between a suppressor effect - due to the fungal requirements of mainly carbon 4